U.S. patent application number 12/484110 was filed with the patent office on 2009-12-17 for assay device and method.
This patent application is currently assigned to AMIC AB. Invention is credited to David BERGMAN, Annika LINDSTROM, lb MENDEL-HARTVIG.
Application Number | 20090311805 12/484110 |
Document ID | / |
Family ID | 41415161 |
Filed Date | 2009-12-17 |
United States Patent
Application |
20090311805 |
Kind Code |
A1 |
BERGMAN; David ; et
al. |
December 17, 2009 |
ASSAY DEVICE AND METHOD
Abstract
There is disclosed an analysis device for the analysis of a
liquid sample, said device comprising a substrate, said substrate
at least partly having projections substantially vertical to the
surface of said substrate, and having a height (H1), diameter (D1)
and center-to-center distance (x1, y1) such, that lateral capillary
flow of said liquid sample is achieved, wherein that said substrate
comprises at least one substrate zone comprising projections
substantially vertical to the surface of said substrate, and having
a height (H2), diameter (D2) and center-to-center distance (x2,
y2), such, that lateral capillary flow of said liquid sample is
achieved and wherein at least one substance is applied at least
partly between the projections in said at least one substrate zone.
Moreover there is provided a method for the analysis of a sample.
The device and method provide for instance improved control of the
dissolution of a substance.
Inventors: |
BERGMAN; David; (Knivsta,
SE) ; MENDEL-HARTVIG; lb; (Uppsala, SE) ;
LINDSTROM; Annika; (Uppsala, SE) |
Correspondence
Address: |
MORRISON & FOERSTER LLP
425 MARKET STREET
SAN FRANCISCO
CA
94105-2482
US
|
Assignee: |
AMIC AB
Uppsala
SE
|
Family ID: |
41415161 |
Appl. No.: |
12/484110 |
Filed: |
June 12, 2009 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
|
61061983 |
Jun 16, 2008 |
|
|
|
Current U.S.
Class: |
436/518 ;
422/68.1 |
Current CPC
Class: |
B01L 3/502746 20130101;
G01N 33/558 20130101 |
Class at
Publication: |
436/518 ;
422/68.1 |
International
Class: |
G01N 33/543 20060101
G01N033/543 |
Claims
1. An analysis device for the analysis of a liquid sample, said
device comprising a substrate, said substrate at least partly
having projections substantially vertical to the surface of said
substrate, and having a height (H1), diameter (D1) and
center-to-center distance (x1, y1) such, that lateral capillary
flow of said liquid sample is achieved, wherein said substrate
comprises at least one substrate zone comprising projections
substantially vertical to the surface of said substrate, and having
a height (H2), diameter (D2) and center-to-center distance (x2,
y2), such, that lateral capillary flow of said liquid sample is
achieved and wherein at least one substance is applied at least
partly between the projections in said at least one substrate
zone.
2. The analysis device according to claim 1, wherein the
projections of the substrate zone(s) have a height (H2), diameter
(D2) and center-to-center distance (x2, y2) such that the capillary
force exerted by the projections facilitate addition of said
substance(s) to the substrate zone(s).
3. The analysis device according to claim 1, wherein the
projections in the substrate zone have a different height (H2),
diameter (D2) or center-to-center distance (x2, y2) so that the
substrate zone has an appearance which is different from the
surrounding.
4. The analysis device according to claim 1, wherein said substrate
further comprises in sequence; at least one sample addition zone,
at least one connecting zone, and at least one receiving zone in
fluid connection, wherein said connecting zone comprises at least
one reaction zone, and wherein said at least one substrate zone is
between said sample addition zone and said reaction zone.
5. The analysis device according to claim 1, wherein at least one
zone is surrounded by an area where at least one of the height (H),
diameter (D) and center-to-center distance (x, y) of the
projections is different so that the surrounding projections exert
a lower capillary force on the sample liquid then the zone.
6. The analysis device according to claim 1, wherein an area
without any projections is surrounding the substrate zone.
7. The analysis device according to claim 4, wherein an area
without any projections is surrounding the reaction zone.
8. The analysis device according to claim 1, wherein said at least
one substance is applied in a volume between the projections in
said at least one substrate zone, so that the substance in a dried
state fills a volume up to essentially the same height (H2) as the
projections.
9. The analysis device according to claim 1, wherein said at least
one substance is applied in a volume between said projections in
said at least one substrate zone, so that the substance in a dried
state fills a volume up to a level so that there is essentially no
liquid flow on top of said at least one substance.
10. The analysis device according to claim 1, wherein the heights
(H1, H2), diameters (D1, D2) and center-to-center distances (x1,
x2, y1, y2) are adapted so that said at least one substance is
gradually dissolved in a flow of liquid flowing by said substrate
zone.
11. The analysis device according to claim 1, further comprising at
least one additional zone, each additional zone comprises
projections substantially vertical to the surface of said
substrate, each with a height (H.sub.n), diameter (D.sub.n) and
center-to-center distance (x.sub.n, y.sub.n), and wherein a
substance is applied in at least one of said at least one
additional zone.
12. The analysis device according to claim 1, wherein the at least
one substrate zone has a shape selected from the group consisting
of a triangle, a square, a rectangle, a parallelogram, a rhombus, a
trapezoid, a quadrilateral, a polygon, a circle, an oval, a half
circle, a half oval, a half polygon, and a circle segment.
13. The analysis device according to claim 1, further comprising a
casing.
14. A method for the analysis of a sample comprising the steps of:
a. addition of a liquid sample on at least one spot on a substrate,
b. performing at least one measurement on the substrate, wherein an
analysis device according to claim 1 is used.
15. The method according to claim 14, wherein the substrate first
is wetted by sample liquid comprising essentially no dissolved
substance and then is brought into contact with liquid sample
comprising dissolved substance.
16. The method according to claim 14, wherein the substrate
comprises a receiving zone with the capacity to receive liquid
sample, and wherein more than 95 wt % of the applied substance is
not dissolved until any part of the liquid sample has reached the
receiving zone.
17. The method according to claim 14, wherein the at least one zone
first comes into contact with liquid sample and when at least a
part of applied substance in said zone has dissolved, the liquid
sample comes into contact with another zone comprising another
applied substance.
Description
[0001] The present application claims the benefit of U.S.
Provisional Application Ser. No. 61/061,983, filed on Jun. 16,
2008, the contents of which are hereby incorporated by reference,
in its entirety.
TECHNICAL FIELD
[0002] The present invention concerns an assay device for the
analysis of a liquid sample.
BACKGROUND
[0003] Quick, reliable, and cost effective analytical and
diagnostic devices for instance devices for use in point of care
are desirable.
[0004] In many assays, detection conjugate and possibly further
reagents are predispensed or integrated in the device, setting
aside the need for separate addition of reagents by the user.
[0005] A common type of disposable assay device comprises a zone
for receiving the sample, a reaction zone, and optionally a
transport or incubation zone connecting the receiving and reaction
zone, respectively. These assay devices are known as
immunochromatography assay devices or simply referred to as strip
tests.
[0006] PCT/SE03/00919 relates to a micro fluidic system comprising
a substrate and provided on said substrate there is at least one
flow path comprising a plurality of micro posts protruding upwards
from said substrate, the spacing between the micro posts being
small enough to induce a capillary action in a liquid sample
applied, so as to force said liquid to move.
[0007] PCT/SE2005/000429 shows a device and method for the
separation of a component in a liquid sample prior to the detection
of an analyte in said sample, wherein a sample is added to a
receiving zone on a substrate, said substrate further optionally
comprising a reaction zone, a transport or incubation zone
connecting the receiving and reaction zone, respectively, forming a
flow path on a substrate, wherein said substrate is a non-porous
substrate, and at least part of said flow path consists of areas of
projections substantially vertical to the surface of said
substrate, and having a height, diameter and reciprocal spacing
such, that lateral capillary flow of said liquid sample in said
zone is achieved, and where means for separation are provided
adjacent to the zone for receiving the sample.
[0008] PCT/SE2005/000787 concerns a device for handling liquid
samples, comprising a flow path with at least one zone for
receiving the sample, and a transport or incubation zone, said
zones connected by or comprising a zone having projections
substantially vertical to its surface, said device provided with a
sink with a capacity of receiving said liquid sample, said sink
comprising a zone having projections substantially vertical to its
surface, and said sink being adapted to respond to an external
influence regulating its capacity to receive said liquid
sample.
[0009] PCT/SE2006/000745 relates to an absorbing zone for
establishing and/or maintaining fluid transport through or along
said at least one fluid passage is manufactured on the basis of a
non-porous substrate, having projections substantially
perpendicular to said surface, and said projections having a
height, diameter and a distance or distances between the
projections such, that lateral capillary flow of said fluid in said
zone is achieved.
[0010] Although the assay devices comprising projections according
to the prior art are working satisfactory there is still room for a
further improvement regarding for instance the control of the
dissolution of a substance that is applied on the device.
SUMMARY OF THE INVENTION
[0011] One object of the present invention is to provide a device
where the control of the dissolution of a predispensed substance is
further improved. There is made available an analysis device for
the analysis of a liquid sample, said device comprising a
substrate, said substrate at least partly having projections
substantially vertical to the surface of said substrate, and having
a height (H1), diameter (D1) and center-to-center distance (x1, y1)
such, that lateral capillary flow of said liquid sample is
achieved, wherein said substrate comprises at least one substrate
zone comprising projections substantially vertical to the surface
of said substrate, and having a height (H2), diameter (D2) and
center-to-center distance (x2, y2), such, that lateral capillary
flow of said liquid sample is achieved and wherein at least one
substance is applied at least partly between the projections in
said at least one substrate zone.
[0012] Further aspects and embodiments of the present invention are
defined in the appended claims which are incorporated herein by
reference.
SHORT DESCRIPTION OF THE DRAWINGS
[0013] The invention will be described in closer detail in the
following description, examples, and attached drawings, in
which
[0014] FIG. 1 shows an embodiment from above, where there is shown
one sample addition zone 1, one connecting zone 2, one receiving
zone 3 as well as a substrate zone 4 comprising projections with a
diameter which is larger than the diameter for the surrounding
projections. There is also a reaction zone 5. Between projections
in the substrate zone 4 there is an applied substance.
[0015] FIG. 2 shows an embodiment where there is a substrate zone
comprising projections with a diameter D2, which is larger than the
diameter D1 for the surrounding projections.
[0016] FIG. 3 shows an embodiment as in FIG. 2 where a substance
has been applied in the substrate zone.
[0017] FIG. 4 shows an embodiment with a substrate zone comprising
projections with a larger diameter than the surrounding
projections. A substance has been applied between the projections
in the substrate zone. The arrows indicate the direction of a flow
of liquid.
[0018] FIG. 5 shows the same embodiment as in FIG. 4 after a period
of time. The arrows indicate that a part of the substance has
dissolved in the liquid and is brought in the direction of the
flow.
[0019] FIG. 6 shows an embodiment with a substrate zone where a
substance has been applied between the projections in the substrate
zone. There is an area surrounding the substrate zone where there
are no projections.
DEFINITIONS
[0020] Before the present device and method is described, it is to
be understood that this invention is not limited to the particular
configurations, method steps, and materials disclosed herein as
such configurations, steps and materials may vary somewhat. It is
also to be understood that the terminology employed herein is used
for the purpose of describing particular embodiments only and is
not intended to be limiting since the scope of the present
invention will be limited only by the appended claims and
equivalents thereof. It must also be noted that, as used in this
specification and the appended claims, the singular forms "a",
"an", and "the" include plural referents unless the context clearly
dictates otherwise. Thus, for example, reference to a reaction
mixture containing "an antibody" includes a mixture of two or more
antibodies.
[0021] The term "about" when used in the context of numeric values
denotes an interval of accuracy, familiar and acceptable to a
person skilled in the art. Said interval can be .+-.10% or
preferably .+-.5%.
[0022] In describing and claiming the device and method, the
following terminology will be used in accordance with the
definitions set out herein.
[0023] As used throughout the claims and the description the term
"analysis" means the process in which at least one analyte is
determined.
[0024] As used throughout the claims and the description the term
"analysis device" means device by the aid of which an analysis can
be performed.
[0025] As used throughout the claims and the description the term
"analyte" means a substance or chemical or biological constituent
of which one or more properties are determined in an analytical
procedure. An analyte or a component itself can often not be
measured, but a measurable property of the analyte can. For
instance, it is possible measure the concentration of an
analyte.
[0026] As used throughout the claims and the description the term
"capillary flow" means flow induced mainly by capillary force.
[0027] As used throughout the claims and the description the term
"casing" means an element enclosing a part of or the entire
substrate.
[0028] As used throughout the claims and the description the term
"center-to-center distance" means the distance between adjacent
projections, measured from the center of a projection to the center
of an adjacent projection. For a planar substrate the
center-to-center distance is measured both in the x direction and
in the y direction, in an orthogonal coordinate system in the
substrate plane.
[0029] As used throughout the claims and the description the term
"center of a projection" means the center of gravity for an
infinitesimal thin slice of the projection taken on half of the
height of the projection in a plane parallel to the surface of the
substrate. For curved substrates the plane is parallel to the
surface of the substrate in a sufficiently small surrounding around
the projection.
[0030] As used throughout the claims and the description the term
"connecting zone" means a zone which establishes fluid connection
between at least two other zones.
[0031] As used throughout the claims and the description the term
"detectable group" means any arrangement of molecules or atoms that
can be detected when present on a substrate.
[0032] As used throughout the claims and the description the term
"fluid connection" means a connection in which a fluid can be
transported.
[0033] As used throughout the claims and the description the term
"sample" means a mixture or a solution to be analyzed.
[0034] As used throughout the claims and the description the term
"substance" means any pure chemical or biological entity or any
mixture or solution comprising at least one chemical or biological
entity.
DETAILED DESCRIPTION
[0035] In a first aspect there is provided an analysis device for
the analysis of a liquid sample, said device comprising a
substrate, said substrate at least partly having projections
substantially vertical to the surface of said substrate, and having
a height, diameter and center-to-center distance such, that lateral
capillary flow of said liquid sample is achieved, and said
substrate comprises at least one substrate zone comprising
projections substantially vertical to the surface of said
substrate, and having a height (H2), diameter (D2) and
center-to-center distance (x2, y2), such, that lateral capillary
flow of said liquid sample is achieved and wherein at least one
substance is applied at least partly between the projections in
said at least one substrate zone.
[0036] When a liquid sample is added to the substrate a flow is
created by capillary forces due to the projections on the surface,
where the projections have a height (H1), diameter (D1) and
center-to-center distance (x1, y1). There is also at least one zone
which is covered with projections, where the projections have a
different height (H2), diameter (D2) or center-to-center distance
(x2, y2), compared to the other projections on the substrate with a
height (H1), diameter (D1) and center-to-center distance (x1, y1).
In one embodiment at least one of the properties height, diameter
and center-to-center distance is different between the zones. In
another embodiment the height, diameter and center-to-center
distance are different. In a further embodiment the diameter and
center-to-center distance are different. In one embodiment H1 and
H2 are different. In one embodiment D1 and D2 are different. In one
embodiment x1 and x2 are different. In one embodiment y1 and y2 are
different. In further embodiments several of the parameters H, D, x
and y are different. In one embodiment all parameters H, D, x and y
are the same for at least two zones of the substance. In one
embodiment all parameters H, D, x and y are the same for the entire
device.
[0037] At least one substance is applied between projections on the
substrate. In one embodiment the substance is applied entirely
within the at least one substrate zone. In an alternative
embodiment at least one substance is applied both in the at least
one substrate zone and outside the at least one substrate zone.
[0038] In one embodiment the projections in the substrate zone have
a different height (H2), diameter (D2) or center-to-center distance
(x2, y2) so that the substrate zone has an appearance which is
different from the surrounding. This is an advantage when the
device is manufactures since the zone where a substance is to be
applied can easily be identified by a human eye or by an automated
device.
[0039] In one embodiment the projections in the substrate zone have
a different height (H2), diameter (D2) or center-to-center distance
(x2, y2) so that the substrate zone exerts a different capillary
force on a substance in solution compared to the surrounding. This
facilitates the addition of a substance in a solution or a
suspension. In one embodiment the capillary force exerted by the
substrate zone is higher than for the surrounding and thus a
substance in a solution or a suspension can easier be applied to
the substrate zone only. The substance to be applied is in one
embodiment dissolved or suspended in a solvent and applied to the
substrate. The solvent is in one embodiment evaporated and the
substance is thus left on the substrate. This has the advantage to
create a well defined area in which the substance is applied. The
boundaries of the applied substance are sharp.
[0040] In one embodiment the substance is applied in a volume
between the projections of the substrate zone. When the substance
is applied to the substrate the capillary force exerted from the
projections of the substrate zone in one embodiment causes the
substance in solution or suspension to fill out essentially the
entire volume between the projections of the substrate zone.
[0041] In one embodiment the projections of the substrate zone(s)
have a height (H2), diameter (D2) and center-to-center distance
(x2, y2) such that the capillary force exerted by the projections
facilitate addition of said substance(s) to the substrate
zone(s).
[0042] In one embodiment the substance is an essentially pure
substance. In another embodiment the at least one substance is a
mixture of two or more substances.
[0043] Examples of substances which can be applied on the substrate
include but are not limited to antibodies, DNA, RNA, aptamers,
antibodies directed to specific analytes in a sample, fragmented
antibodies, antibody fragments, synthetic binders, chemical
binders, receptors, ligands, affibodies, cells, organelles,
polypeptides, peptides, enzymes, monoclonal antibodies, polyclonal
antibodies, phage display proteins, IgG immunoglobulins, chemical
ligands, and combinations thereof.
[0044] In one embodiment the substance to be applied to the
substrate comprises at least one further additive. Examples of
further additives include but are not limited to sugars, polymers,
detergents, surface active agents, cationic surface active agents,
non-ionic surface active agents, anionic surface active agents,
salts, and lipids or any combination thereof.
[0045] In one embodiment there is added at least one further
substance to the substrate, where that substance is at least one
substance selected from a sugar, a polymer, a detergent, a surface
active agent, a cationic surface active agent, a non-ionic surface
active agent, an anionic surface active agent, a salt, and a
lipid.
[0046] In one embodiment the substrate further comprises in
sequence; at least one sample addition zone, at least one
connecting zone, and at least one receiving zone in fluid
connection, wherein said connecting zone comprises at least one
reaction zone, and wherein said at least one substrate zone is
between said sample addition zone and said reaction zone. In one
embodiment at least a part of the connecting zone is covered by
projections such that a lateral capillary flow is achieved.
[0047] In one embodiment at least one zone is surrounded by an area
where at least one of the height (H), diameter (D) and
center-to-center distance (x, y) of the projections is different so
that the surrounding projections exert a lower capillary force on
the sample liquid then the zone. In one embodiment said zone is at
least one zone selected from the reaction zone and the substrate
zone. In one embodiment said zone is the substrate zone. In one
embodiment the center-to-center distance in an area surrounding a
zone is larger compared to within the zone.
[0048] In one embodiment there is an area without any projections
surrounding at least one of the substrate zone and the reaction
zone. In FIG. 6 there is depicted one embodiment where an area
without projections is surrounding a substrate zone. In one
embodiment the surrounding area without projections is from 15 to
100 .mu.m wide. In another embodiment the surrounding area without
projections is from 20 to 40 .mu.m wide. In a further embodiment
the surrounding area without projections is from 25 to 35 .mu.m
wide. The width depends on the viscosity of the substrate which is
added to the substrate zone and on the hydrophilicity of the
surface surrounding the substrate zone.
[0049] Advantages of a zone with a surrounding area with different
properties include that it is possible to apply a substance to the
zone without the substance flowing out of the zone. Thus it is
possible to apply a substance to a zone in a reproducible and well
defined manner.
[0050] In one embodiment the entire connecting zone is covered by
projections such that a lateral capillary flow is achieved. In one
embodiment at least a part of the sample addition zone, the
connecting zone and the receiving zone are covered by projections
such that a lateral capillary flow is achieved. In another
embodiment the entire sample addition zone, connecting zone and
receiving zone are covered by projections such that a lateral
capillary flow is achieved.
[0051] In one embodiment the analysis device and the substrate is
such that it is possible to add a sample to the sample addition
zone. Due to capillary flow induced by the projections a lateral
flow is created and at least a part of the sample reaches at least
one substrate zone on the substrate where there are projections
with a height (H2), diameter (D2) and center-to-center distance
(x2, y2). In one embodiment the substance applied in the at least
one substrate zone and/or near the at least one substrate zone is
gradually dissolved by the sample flowing by the at least one
substrate zone. The liquid sample thereafter reaches the reaction
zone. In one embodiment a measurement is made in the reaction zone.
Examples of measurement techniques include but are not limited to
detection of fluorescence, and chemiluminescence. A person skilled
in the art realizes that also other detection principles can be
used such as absorption of light optionally at several different
wavelengths, and detection of emitted light. In one embodiment the
sample continues to a receiving zone, which receives the liquid
sample. The flow continues until there is no more liquid sample or
until the receiving zone is full of sample liquid.
[0052] In one embodiment there is a surplus of sample liquid. In
one embodiment a part of the surplus of sample liquid is used so
that when all of the substance has been dissolved the reaction zone
and other parts of the device is washed so that free or loosely
bound substance is washed away.
[0053] In one embodiment there are several receiving zones.
[0054] The at least one substance, which is applied to the at least
one substrate zone, is in one embodiment applied to the substrate
surface. In one embodiment the substance is applied between the
projections. In one embodiment the thickness of the applied
substance in a dried state is essentially corresponding to the
height of the projections. In an alternative embodiment the
thickness of the applied substance in a dried state is lower than
the height of the projections. In one embodiment the thickness of
the applied substance in a dried state is higher than the
projections.
[0055] In one embodiment the at least one substance is applied in a
volume between the projections in said at least one substrate zone,
so that the substance in a dried state fills a volume up to a level
at half of the height (H2) of the projections.
[0056] In an alternative embodiment the at least one substance is
applied in a volume between the projections in said at least one
substrate zone, so that the substance in a dried state fills a
volume up to essentially the same height (H2) as the
projections.
[0057] In a further embodiment the at least one substance is
applied in a volume between said projections in said at least one
substrate zone, so that the substance in a dried state fills a
volume up to a level so that there is essentially no liquid flow on
top of said at least one substance.
[0058] In one embodiment the volume between the projections of the
substrate zone is adjusted so that the amount of substance which is
to be applied fills up to the desired level in a dried state.
[0059] In one embodiment the substance is applied in a solution and
is dried so that a solvent is evaporated. In one embodiment the
solvent is water. When the solvent has evaporated the remaining
substance is called the substance in a dried state.
[0060] In embodiments where the there is no flow or essentially no
flow of liquid on top of the applied substance, the dissolution of
the applied substance occurs from the sides and not from the top.
This gives a controlled dissolution. The dissolution occurs during
a prolonged time and in a more controlled manner compared to an
embodiment where the liquid sample also flows over the applied
substance.
[0061] In one embodiment no liquid flows on top of the applied
substance. In an alternative embodiment only a minor part of the
liquid flows on top of the applied substance, examples of such a
part of the liquid include but are not limited to 0.1 wt %, 1 wt %,
5 wt %, and 10 wt %. In one embodiment less than 1 wt % of the
liquid flows on top of the applied substance. In another embodiment
less than 10 wt % of the liquid flows on top of the applied
substance.
[0062] By adjusting the diameter and center-to-center distance of
the projections for instance in an embodiment where essentially no
liquid flows over the applied substance, it is possible to control
the dissolution of the applied substance in the liquid.
[0063] In one embodiment all components of the applied substance
are dissolved by the liquid sample. In an alternative embodiment
not all applied substances are dissolved by the liquid sample.
[0064] By adjusting the height, diameter and center-to-center
distance of the projections in the zone where a substance has been
applied and outside the zone where a substance has been applied it
is possible to control the lateral capillary flow so that a desired
dissolution of the applied substance occurs at a desired rate.
[0065] In one embodiment the heights (H1, H2), diameters (D1, D2)
and center-to-center distances (x1, x2, y1, y2) are adapted so that
said at least one substance is gradually dissolved in a flow of a
liquid flowing by said substrate zone.
[0066] The invention is not limited to two different heights,
diameters and center-to-center distances of the projections. There
are provided embodiments with further zones, where each zone has a
height, diameter, and center-to-center distance, where at least one
of the parameters is different compared to the other zones.
[0067] There is provided an embodiment where the analysis device
comprises at least one additional zone n, where each additional
zone n comprises projections substantially vertical to the surface
of said substrate, with a height (H.sub.n), diameter (D.sub.n) and
center-to-center distance (x.sub.n, y.sub.n), such that lateral
capillary flow is created. There is provided a substrate having n
distinct zones, each zone comprising projections having a height
(H.sub.n), diameter (D.sub.n) and center-to-center distance
(x.sub.n, y.sub.n), where n is a natural number. n=1, 2, 3, 4, 5, 6
. . .
[0068] In one embodiment there is at least one substance applied in
at least one of the n zones.
[0069] In one embodiment there is a substrate zone comprising
projections substantially vertical to the surface of said
substrate, and having a height, diameter and center-to-center
distance, such, that lateral capillary flow of said liquid sample
is achieved, where the height, diameter and center-to-center
distance in the substrate zone is different from outside of the
substrate zone. In one embodiment the height, diameter and
center-to-center distance of the projections vary within the
substrate zone.
[0070] In one embodiment there are at least two zones comprising an
applied substance, which zones have the shape of concentric
circles. In one embodiment the outer part comprises at least one
applied substance and the inner part comprises at least one another
applied substance. First the outer substance is dissolved and then
the inner substance is dissolved. Thereby further possibilities to
control the dissolution are provided. There is also provided
possibility to dissolve several substance in sequence or together.
In one embodiment there is provided the possibility to dissolve
more than one substance in sequence. In other embodiments also
other shapes of the different zones with applied substances are
provided. In one embodiment there is at least one zone which first
comes into contact with the sample liquid and when a fraction of or
all of the substance in that zone has dissolved the sample liquid
comes into contact with another zone comprising a different
substance. Thereby it is possible to start the dissolution of a
substance in the sample liquid after a certain period of time.
[0071] Thus there is provided a method wherein the at least one
zone first comes into contact with liquid sample and when at least
a part of applied substance in said zone has dissolved, the liquid
sample comes into contact with another zone comprising another
applied substance.
[0072] In one embodiment there are two different zones, where each
zone comprises a distinct substance.
[0073] In one embodiment a part of the liquid sample passes the
zone(s) where a substance has been applied at such a distance that
essentially no substance is dissolved, and another part passes near
the zone where a substance has been applied so that the substance
dissolves in the liquid. Thus there is a provided an embodiment
where a part of the liquid stream flowing through the device
comprises a major part of dissolved substance and another part
comprises little or no dissolved substance. Examples of a major
part of the dissolved substance include but are not limited to 75
wt %, 90 wt %, 95 wt %, 99 wt %, and 99.9 wt %.
[0074] It is possible to control the height, diameter and
center-to-center distance of the projections inside and outside of
the zone where the substance to be dissolved is applied. In that
way it is possible to control the fraction of the liquid sample
that flows by the zone where the substance is dissolved. By
controlling the rate at which the substance dissolves when the
liquid sample flows by, it is possible to control the concentration
of the dissolved substance.
[0075] If a larger fraction of liquid sample passes outside the
zone where the substance is dissolved at a high flow, the result is
that downstream of the zone where the sample is applied; there is a
rather narrow concentrated flow of dissolved substance. On the
other hand, if a smaller fraction of liquid sample passes the zone
where the substance is dissolved the result is that downstream of
the zone where the sample is applied; there is a broader trace of
dissolve substance. The trace of dissolved substance is shown in
FIG. 5 as a grey area.
[0076] In one embodiment the result is detected in the middle of
the trace of dissolved substance downstream of the zone where the
substance is applied. In one embodiment the result of the analysis
is read in the reaction zone, downstream of the zone where the
substance is applied.
[0077] The shape of the zone where the substance is applied is in
one embodiment adapted to control the dissolution rate and/or how
the dissolved substance is distributed in the flow of liquid
sample.
[0078] Examples of shapes include but are not limited to a
triangle, a square, a rectangle, a parallelogram, a rhombus, a
trapezoid, a quadrilateral, a polygon, a circle, and an oval.
[0079] Also truncated shapes are encompassed, including but not
limited to a half circle, a half oval, a half polygon, and a circle
segment.
[0080] Further shapes include all possible combinations of shapes
including but not limited to a triangle and a square, a triangle
and a rectangle, a trapezoid and a rectangle, a half circle and a
rectangle and so on.
[0081] In one embodiment the shape of the zone where the substance
is applied is triangular with one of the corners pointing in the
direction from where the flow of sample liquid comes.
[0082] In one embodiment the shape of the zone where the substance
is applied is an isosceles trapezoid.
[0083] In one embodiment the shape of the zone where the substance
is applied is an isosceles trapezoid combined with a rectangle.
[0084] In one embodiment the shape of the zone where the substance
is applied is an isosceles trapezoid combined with a rectangle,
with the narrower zone pointing towards the direction from where
the flow of sample liquid comes.
[0085] In one embodiment the shape of the zone where is substance
is applied and the height, diameter and center-to-center distance
of the projections are adapted so that the concentration of the
dissolved substance is greater towards the middle of the
substrate.
[0086] In one embodiment the analysis device further comprises a
lid. Preferably the lid has at least one opening or an aperture for
the addition of a liquid sample. In one embodiment there is an
opening or a window allowing a measurement result to be read from
the analysis device. If a lid is used the lid is not in capillary
contact with the projections on the substrate. The lid does not
take part in creating any capillary forces.
[0087] In one embodiment the analysis device comprises a casing. In
one embodiment the casing encloses the entire or a part of the
analysis substrate. Preferably the case has at least one opening or
an aperture for the addition of a liquid sample. In one embodiment
there is an opening or a window allowing a measurement result to be
read from the analysis device.
[0088] In one embodiment the applied substance comprises a
detection conjugate. In one embodiment the detection conjugate
comprises at least one element selected from an antibody, DNA, RNA,
an aptamer, a fragmented antibody, an antibody fragment, a
synthetic binder, a chemical binder, a receptor, a ligand, an
affibody, a cell, an organelle, a polypeptide, a peptide, an
enzyme, a monoclonal antibody, a polyclonal antibody, a phage
display protein, an IgG immunoglobulin, a chemical ligand. In one
embodiment the detection conjugate comprises more than one
antibody. In one embodiment at least one of the molecules in the
applied substance comprises a detectable group, which allows
detection for instance in the reaction zone. The detection
conjugate facilitates detection of an antigen bound to the
conjugate. In one embodiment the detection conjugate comprises a
fluorescent molecule. In one embodiment fluorescence from the
detection conjugate is measured.
[0089] In one embodiment at least one selected from an antibody,
DNA, RNA, an aptamer, a fragmented antibody, an antibody fragment,
a synthetic binder, a chemical binder, a receptor, a ligand, an
affibody, a cell, an organelle, a polypeptide, a peptide, an
enzyme, a monoclonal antibody, a polyclonal antibody, a phage
display protein, an IgG immunoglobulin, and a chemical ligand, is
bound to the substrate downstream of the applied substance. Such an
antibody or aptamer is able to bind to the complex between an
antibody and an antigen or to a free antigen. In one embodiment an
antibody or an aptamer bound to the reaction zone is able to bind
to the complex between an antibody and an antigen or to a free
antigen. In one embodiment fluorescence from the detection
conjugate in the reaction zone is measured.
[0090] In a second aspect there is provided a method for the
analysis of a sample comprising the steps of: a) addition of a
liquid sample on at least one spot on a substrate, and b)
performing at least one measurement on the substrate, wherein an
analysis device as described herein is used.
[0091] In one embodiment the substrate is first wetted by sample
liquid comprising essentially no dissolved substance and then is
brought into contact with liquid sample comprising dissolved
substance. The first wetting by the liquid sample is achieved by a
delay of the dissolution of the substance applied to the substrate.
The sample liquid that flows first comprises essentially no
dissolved substance, such as but not limited to less than 0.001 wt
%, 0.01 wt %, 0.1 wt % or 1 wt %. When the liquid sample comes into
contact with the substance, the dissolution process starts and the
level of dissolved substance gradually increases.
[0092] In one embodiment there are molecules bound to the substrate
which are first hydrated by the liquid sample, where the liquid
sample is essentially without any dissolved substance. In one
embodiment such molecules are antibodies. One advantage of such a
pre-hydration is that the antibodies or molecules become more
active. This first hydration is called a pre wetting. In one
embodiment a substantial part of the substance(s) applied to the
substrate is dissolved and transported across the reaction zone
after a steady lateral flow has been established by the capillary
force from the receiving zone. A substantial part of the
substance(s) applied to the substrate is in this case more than 75
wt %, preferably more than 90 wt %, more preferably more than 95 wt
% and most preferably more than 99 wt %.
[0093] In one embodiment the sample is added to a sample addition
zone, the sample flows through a connecting zone to a receiving
zone. The receiving zone has a capacity to receive the sample
liquid and has a large surplus of capillary force. The capillary
force of the receiving zone is in one embodiment such that a steady
and even lateral flow of sample liquid is created. The capillary
force of the receiving zone acts like a pump and receives the
sample liquid at a steady rate. Before the added sample liquid has
reached the receiving zone the lateral capillary flow is not always
even and steady.
[0094] It is one advantage that a substantial part of the applied
substance is not dissolved until there is a steady lateral flow,
i.e. when the liquid sample has reached the receiving zone. In that
way the dissolution of the substance occurs at a more controlled
way due to the steady and even flow of sample liquid.
[0095] In one embodiment the substrate comprises a receiving zone
with the capacity to receive liquid sample, and wherein more than
95 wt % of the applied substance is not dissolved until any part of
the liquid sample has reached the receiving zone. In an alternative
embodiment more than 90 wt %, preferably 94 wt %, more preferably
99 wt % and most preferably 99.9 wt % of the applied substance is
not dissolved until any part of the liquid sample has reached the
receiving zone. This implies that at most 10 wt %, preferably 6 wt
%, more preferably 1 wt % and most preferably 0.1 wt % of the
applied substance is dissolved until any part of the liquid sample
reaches the receiving zone.
[0096] Further advantages include that there is provided a
possibility of increased control of the flow. There is also
provided the possibility of dissolution during a steady lateral
flow. It is possible to control the start and stop of dissolution
of a substance. There is a possibility to provide a wash of the
reaction zone after the dissolved substance(s) has passed. It is
possible to obtain an even dissolution of the reagent over time.
There is the possibility to ensure that the dissolved substance is
spatially homogenously distributed. There is also provided the
possibility to control the spatial distribution of the dissolved
substance. It is possible to ensure that all substance is dissolved
in the sample. A major portion of the sample is contacted with the
substance.
EXAMPLES
Example 1
[0097] A substrate with dimensions 25.times.75 mm was made by
injection molding of a cyclo olefin polymer (COP). The substrate
hade a sample addition zone, a substrate zone, a connecting zone, a
reaction zone, and a receiving zone. On the substrate there were
projections with height 70 .mu.m and diameter 50 .mu.m. The
distance between the projections were 15 .mu.m in the sample
addition zone, the substrate zone, the connecting zone, and the
reaction zone. A distance between the projections of 15 .mu.m and a
diameter of 50 .mu.m corresponds to a center-to-center distance of
65 .mu.m between the projections in both x- and y-direction. The
substance zone was surrounded by a gap where the distance between
the projections was 30 .mu.m.
[0098] It was possible to apply detection conjugate to the
substance zone so that the detection conjugate did not move out of
the substrate zone. The application of the detection conjugate was
reproducible during several applications. The detection conjugate
was applied so that the height of the dried detection conjugate was
at about the same level as the top of the projections in the
substance zone.
[0099] A detection conjugate comprising antibodies against
N-terminal pro-brain natriuretic peptide (NT-proBNP) was used.
[0100] Separated whole blood was added to the sample addition zone
and the detection conjugate dissolved gradually in the liquid
sample. The trace of dissolved detection conjugate was continuous
and the dissolution time could be controlled. The experiment was
repeated six times and the dissolution time of the detection
conjugate was 6 minutes and 8 seconds, whereby the coefficient of
variation was 3%.
[0101] The total assay time was 19 minutes and 19 seconds with a
coefficient of variation of 5% (six repeated experiments).
[0102] The result of the analysis was read by using an optical
reader.
[0103] Although the invention has been described with regard to its
preferred embodiments, which constitute the best mode presently
known to the inventors, it should be understood that various
changes and modifications as would be obvious to one having the
ordinary skill in this art may be made without departing from the
scope of the invention which is set forth in the claims appended
hereto.
* * * * *